Helicobacter cholecystus Franklin et al.
700242 ™
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700242 ™
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ATCC highly recommends that appropriate personal protective equipment is always used when handling vials. For cultures that require storage in liquid nitrogen, it is important to note that some vials may leak when submersed in liquid nitrogen and will slowly fill with liquid nitrogen. Upon thawing, the conversion of the liquid nitrogen back to its gas phase may result in the vial exploding or blowing off its cap with dangerous force creating flying debris. Unless necessary, ATCC recommends that these cultures be stored in the vapor phase of liquid nitrogen rather than submersed in liquid nitrogen.
2. Aseptically withdraw the contents and add to a tube of #18 Tryptic Soy Broth.
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3. To obtain a biphasic culture, add 0.5 ml of the suspension to a #260 slant. Add remaining 0.1 ml of the suspension to a #260 plate and streak for isolation.
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4. Incubate at 37°C under microaerophilic conditions using an anaerobe jar with an active catalyst and a microaerophilic gas generator pack, or other acceptable method, to obtain microaerophilic conditions. Incubate slant with cap loose.
5. Within three days of incubation, growth should be obtained in the broth pool at the bottom of the slant. Additional incubation may be required for colonies to appear on the plate. Further subcultures can be made using broth pool as the inoculum source.
5.
This is a slow growing organism that requires moist conditions for best growth. Growth at the broth/agar interface of the biphasic slant should occur within three days, but little turbidity will be seen. To observe growth, examine a wet mount of the broth under phase microscopy. The organism is a small slightly curved rod with many coccoid forms, both exhibit motility. Motility is usually observed only in young cultures. The presence of spheroid cells indicates that viability is being lost either due to age or too much exposure to oxygen.
Growth on agar takes longer than with the biphasic culture. Colonies are small, circular, entire, convex, smooth, and gray. Once good growth is present, these organisms tend to lose viability, especially if exposed to air for lengthy periods. Viability also decreases with repeated subculturing. The cells do not Gram stain well using traditional procedures. To obtain the best results, use a basic fuchsin counterstain in place of the safranin.
Once good growth is obtained, transfer or freeze the culture. Adding an equal amount of 20% sterile glycerol to pooled broth from several biphasic slants, followed by freezing in liquid nitrogen or "ultra-low temperature" freezer is recommended.
Additional information on this culture is available on the ATCC web site at www.atcc.org.
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Validation list no. 61. Int. J. Syst. Bacteriol. 47: 601-602, 1997.
Franklin CL, et al. Isolation of a novel Helicobacter species, Helicobacter cholecystus sp. nov., from the gallbladders of syrian hamsters with cholangiofibrosis and centrilobular pancreatitis. J. Clin. Microbiol. 34: 2952-2958, 1996. PubMed: 8940429
type strain